The introduction of the polymerase chain reaction (PCR) and subsequent alternative amplification systems for nucleic acids has enabled the use of this genetic material as a specimen for diagnostic tests. As a consequence new analytical methods are now available especially for diagnosing hereditary diseases, a predisposition for certain diseases and infectious diseases which, among others, enable an early diagnosis of the condition.
In order to convert the genetic material into a suitable form for enzymatic amplification, it is necessary to release it from the biological sample material. In addition the nucleic acid must be protected from degradation by nucleases from the biological material or the environment and from degradation by chemical reaction conditions. The most stringent requirements concern the freedom from contamination of the biological sample and the nucleic acid isolated therefrom. For the amplification the nucleic acid should be present in a buffered, aqueous, substantially salt-free solution.
Whereas PCR usually uses very small amounts of analyte (pg-ng range), special problems require the processing of a larger quantity of sample. For example, in order to identify circulating tumour cells at a sensitivity of one tumour cell against a background of normal cells, the nucleic acid has to be isolated from 10-20 ml of a blood sample. After homogenizing the sample, an aliquot of the isolated RNA can then be examined for the expression of a tumour-associated gene.
In addition to the classical methods of nucleic acid isolation by means of enzymatic, mechanical or chemical lysis of the sample material, subsequent extraction of the proteins and lipids of phenol and phenol/CHCl3 and precipitation of the nucleic acid from the aqueous phase by ethanol or i-propanol (Sambrook, J., et al., Molecular Cloning, Cold Spring Harbor Laboratory Press, 1989, 2nd edition, 9.16-9.23; Ausubel, F. M., et al., Current Protocols in Molecular Biology, John Wiley & Sons, 1987, 2.1.1-2.4.5), several commercial kits have been developed especially for PCR sample preparation in recent years which utilize the property of nucleic acids to bind to glass surfaces under chaotropic salt conditions which has been known since the end of the seventies (Vogelstein, B., et al., Proc. Natl. Acad. Sci. USA 75 (1979) 615-619). Other constituents of biological material such as proteins, lipids or salts, are not bound and are therefore separated. Centrifugation vessels with glass fleece inlays or silica gel suspensions which allow a batch process are known. In addition multiple devices are known in a strip and 96-well microtitre plate format with the glass fleeces recessed into the base which can be operated with the aid of a vacuum chamber attached underneath as well as by centrifugation. In these methods the volume of the samples is often limited. Moreover, large amounts of buffer are necessary to effectively elute the nucleic acids from the glass fleeces which results in a diluted solution of the isolated molecules and requires additional preparation steps for certain applications.
A modified method (Miller et al., Nucl. Acids Res. (1988) 16: 1215) uses a concentrated salt solution to precipitate proteins and other accompanying substances after lysis of the sample material. The nucleic acids in the supernatant are then precipitated by ethanol and collected by centrifugation. After the nucleic acids have been dissolved they can be used for amplification.
WO 93/11221 discloses a method and a device for isolating and purifying nucleic acids which uses anion exchangers and mineral carrier substances. U.S. Pat. No. 5,104,533 discloses a filtration unit with pressure compensation. U.S. Pat. No. 4,270,921 discloses the combination of a microcolumn and a centrifuge tube. WO 98/32877 discloses a device for isolating nucleic acids which is composed of two vessels connected by a closure element which contains a material for binding nucleic acids. U.S. Pat. No. 4,956,298 discloses a separation or reaction column consisting of a centrifuging vessel and a receiving body where the receiving body contains a column material and the centrifuging vessel collects the effluent of the receiving body. DE 19512361 discloses a method for isolating a biological material which uses a compressible porous matrix to bind the biological material and compresses the material in order to elute the material. EP 588564 describes a device for affinity separation comprising a capture membrane located in a pipette tip. WO 96/41810 discloses the removal of DNA from a cell suspension with the aid of a hollow membrane filter and an ion exchange step. The manufacture of a device containing a material for binding nucleic acids is known from EP 738733. WO 02/053256 discloses a device and a method for purification comprising a sample holder with a column insert portion, whereas a column module is secured in said column insert portion. U.S. Pat. No. 6,177,009 and the German utility models DE 298 03 712 U1 and DE 202 18 503 U1 describe a device for treating biomolecules comprising a separation column which has a separation device and a collecting vessel for the liquid that flows out.